Use of ambroxol to improve skin barrier function

ABSTRACT

Compositions and methods for improving skin health and skin barrier function. Specifically, topical administration of compositions comprising low dose formulations of ambroxol can be used to improve skin barrier function.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a Continuation Application of U.S. Ser. No.16/595,621, filed on Oct. 8, 2019, which is a Continuation Applicationof PCT/US19/15137 filed on Jan. 25, 2019, which claims priority to U.S.62/622,375 filed on Jan. 26, 2018, all of which are hereby incorporatedby reference in their entirety.

BACKGROUND OF THE INVENTION Field of the Invention

The invention relates to the use of ambroxol to improve skin barrierfunction in a subject, preferably a human subject. Examples of such usesinclude increasing skin hydration, cosmetic uses, reduction of burns,scars, and/or rashes. The invention further relates to inhibiting and/orreducing transepidermal water loss (TEWL) and/or increasing the amountand/or concentration of ceramides (CER) in the skin's epidermis and/orhair by the topical application of ambroxol.

Discussion of the Related Art

In the following discussion, certain articles and methods will bedescribed for background and introductory purposes. Nothing containedherein is to be construed as an “admission” of prior art. Applicantexpressly reserves the right to demonstrate, where appropriate, that thearticles and methods referenced herein do not constitute prior art underthe applicable statutory provisions.

The major function of the skin is to form a barrier between the internalmilieu of a host and environmental insults, such as chemicals,ultraviolet light, mechanical insults and pathogenic microorganisms. Theskin's structure is made up of layers of cells and cellular-derivedtissue, with the outermost set of layers being the epidermis. Thesurface layer is called the stratum corneum then, proceeding inwards:the stratum granulosum, stratum spinosum, stratum basale, and then thedermis layer. In addition, the palms of the hands and the soles of thefeet have an additional layer, the stratum lucidum, between the stratumcorneum and the stratum granulosum. Numerous products have beendeveloped to improve the functioning and/or appearance of skin and hair.

For example, in 2016, the beauty and personal care market was close to a500-billion-dollar industry. Repeated exposure to environmental insults,such as the sun, causes skin thinning, fragility and wrinkles. Moreserious sun exposure can cause a loss of skin elasticity, deep wrinkles,increased roughness and dryness, and altered pigmentation (age spots,skin spots). The skin can also become leathery, thickened in appearancewhich is characterized by deep furrows. Other environmental insults,such as chemicals causing burns and/or injury, acne or viral infection(e.g., chicken pox) can cause scaring, resulting in more serious andpermanent damage to the skin.

Additionally, diseases can also cause changes in the skin's appearance.For example, Lupus patients often exhibit a characteristic rash, terms a“butterfly rash” that spans the face of the patient.

Additionally, the robustness and integrity of the skin, particularlywith respect to its ability to respond to environmental stresses and/orresist disease, generally declines with age.

There are numerous different skin care formulations that are marketed asimproving the appearance of the skin regardless of the type of skininsult. Examples of such formulations include, but are not limited toretinols, hydroxyl acids, Coenzyme Q10, copper peptides, Kinetin, and/ortea extracts. Even though numerous creams, oils, topicals and oralproducts are launched every month, there is an ongoing need for newregimens that are safe and effective in improving the functioning of asubject's largest organ—the skin.

SUMMARY OF THE INVENTION

This Summary is provided to introduce a selection of concepts in asimplified form that are further described below in the DetailedDescription. This Summary is not intended to identify key or essentialfeatures of the claimed subject matter, nor is it intended to be used tolimit the scope of the claimed subject matter. Other features, details,utilities, and advantages of the claimed subject matter will be apparentfrom the following written Detailed Description including those aspectsillustrated in the accompanying drawings and defined in the appendedclaims.

The invention relates to ambroxol compositions and use thereof toimprove skin barrier function. Specifically, the invention relates toimproving skin barrier function of a subject, preferably a human,comprising administering to the subject an effective amount of ambroxol.Ambroxol can be administered as a composition with other components fortopical delivery to a subject's skin and/or hair. In preferredembodiments, a topical composition comprising a low dose formulation ofambroxol is used to improve skin barrier function. In further preferredembodiments, skin barrier function is improved by up to about 10%, 20%,30%, 40%, 50%, 60%, or 70% as compared to untreated control subjects.

A skin's barrier function can be improved with topical administration ofambroxol as measured by:

-   -   a. quickening the recovery of the skin barrier function after an        insult;    -   b. decreasing the delay in the skin barrier recovery, as often        occurs in premature infants or the elderly;    -   c. decreasing/preventing transepidermal water loss (TEWL);    -   d. decreasing/preventing electrolyte loss in the skin;    -   e. decreasing/preventing dry skin;    -   f. decreasing/preventing pruritus;    -   g. decreasing/healing/preventing skin lesions;    -   h. increasing/improving skin hydration;    -   i. improving protection from foreign insults (e.g.,        antimicrobial barrier);    -   j. increasing ceramide (CER) and/or glucosyl-ceramide        production, preferably in the stratum basale, stratum spinosum,        stratum granulosum, and/or stratum corneum layers of the skin;    -   k. increasing conversion of glucosyl-CERs into CER, preferably        in the stratum granulosum and stratum corneum layers of the        skin;    -   l. increasing the amount and/or concentration of CER in the        skin, preferably in the stratum corneum layer of the skin;    -   m. increasing glucocerebrosidase activity in the stratum basale,        stratum spinosum, stratum granulosum, and/or stratum corneum        layers of the skin; or    -   n. decreasing the overall rate of loss of robustness of the skin        upon aging, according to generally accepted functional metrics        that monitor such a decline,    -   wherein the percentage change (i.e., increase or decrease) of        (a)-(n) is at least 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%,        55%, 60%, or 65% as compared to the untreated state.

In a further preferred embodiment, topical formulations of ambroxol asdescribed herein can be used to improve overall appearance of the skin,including treating, reversing and/or preventing symptoms of skin agingor damage, such as wrinkles, fine lines, discoloration, loss of tone,loss of elasticity, thinning and the like, by activating autophagywithin skin cells.

Compositions of ambroxol formulations are provided in a variety of formssuitable for topical application, such as cosmetics, lotions, creams,shampoos, serums, sprays, skin lotions, and cleansers. Similarly,ambroxol formulations are provided in a kit comprising a cleanser, aspray essence, a lotion, a serum and an anti-aging night cream.

Additionally, ambroxol formulations can be used to help remove toxins atthe cellular level, as well as helping boost skin's integrity to:

-   -   a. prevent cellular damage;    -   b. repair cellular damage;    -   c. rebuild and repair the skin's structural network;    -   d. reinvigorate skin cell function; and/or    -   e. repair, remove and/or recycle damaged cellular components for        healthier functioning cells.

In further preferred embodiments, topical formulations of ambroxol asdescribed herein can be used to induce autophagy within the skin cells.

In preferred embodiments, the concentration of CER that is increased inthe stratum corneum after topical administration of ambroxol is at least10%, 20%, 30%, 40%, 50% or 60% higher as when compared to theconcentrations of cholesterol or free fatty acids or other suitablebenchmarks.

By improving the skin barrier function, the appearance of the skinand/or hair is also improved. Such improvements include but are notlimited to reducing the signs of aging (or premature aging), such as forexample, decreasing the aging process of the skin, reducing theappearance of lines (e.g., fine lines) and/or wrinkles, decreasing skinsagging and/or imperfections, and/or improving and/or promoting skinelasticity or suppleness, and/or increasing and/or promoting retentionof fluid in the skin. In other embodiments, the appearance of the skinand/or hair includes but is not limited to, reducing sun damage(particularly UV radiation-induced oxidative stress/photodamage),stretch marks, and/or scars, reducing the appearance of burns, reducingthe appearance of blemishes (e.g. acne), reducing the appearance of darkcircles, age spots, and other hyperpigmentation, reducing skinthickness, loss of elasticity and/or collagen content, reducing dry skinand/or puffiness, preventing water loss of the skin, and/or increasingmoisturization, and/or reducing lentigines, freckles, and/or melasmas.Topical administration also includes the application of ambroxol tohair.

Many medications, cosmetics and inflammatory skin diseases, in additionto adverse reactions to chemical peels, ingredients, pesticides,chemicals, detergents, heat, laser resurfacing, laser assisted hairremoval and skin injuries, can also be a source of pigmentationdisorders. These pigmentation disorders can also be treated by ambroxolas described herein.

Other uses of topical administration of ambroxol as described hereininclude, but are not limited to, treatment of: atopic dermatitis,psoriasis; patients using topical tretinoin, premature infants, burns,and the elderly.

In some embodiments, enantiomers, analogs, esters, amides, prodrugs, ormetabolites of ambroxol, including salts thereof and particularly apharmaceutically acceptable salt can be used. In preferred embodiments,the salt of ambroxol is a hydrochloride.

In preferred embodiments, the topical administration of ambroxol occursat a low dose formulation. As used herein a “low dose formulation”comprises a concentration of ambroxol of less than 0.1% (w/w), and morepreferably, less than 0.01% (w/w), and even more preferably between0.01%-0.09% (w/w), and even more preferably between 0.001%-0.01% (w/w),and even more preferably between 0.0001-0.001% (w/w), and even morepreferably between 0.00001-0.0001% (w/w). In further preferredembodiments, the low dose formulation concentration of ambroxol fortopical administration occurs at about 0.00001% (w/w), 0.00002% (w/w),0.00003% (w/w), 0.00004% (w/w), 0.00005% (w/w), 0.00006% (w/w), 0.00007%(w/w), 0.00008% (w/w), 0.00009% (w/w), 0.0001% (w/w), 0.0002% (w/w),0.0003% (w/w), 0.0004% (w/w), 0.0005% (w/w), 0.0006% (w/w), 0.0007%(w/w), 0.0008% (w/w), 0.0009% (w/w), 0.001% (w/w), 0.002% (w/w), 0.003%(w/w), 0.004% (w/w), 0.005% (w/w), 0.006% (w/w), 0.007% (w/w), 0.008%(w/w), 0.009% (w/w), 0.01% (w/w), 0.02% (w/w), 0.03% (w/w), 0.04% (w/w),0.05% (w/w), 0.06% (w/w), 0.07% (w/w), 0.08% (w/w), or 0.09% (w/w), aswell as any combination of these low dose formulation concentrations. Itis unexpected and not suggested in the literature that ambroxol couldimprove skin barrier function at such low formulation concentrations.

Ambroxol can be formulated with other compounds for topicaladministration as described herein. For example, ambroxol formulationscan include ceramides. Preferred examples of such ceramides include butare not limited to: Ceramide 1 (EOS), Ceramide 2 (NS), Ceramide 3 (NP),Ceramide 4 (EOH), Ceramide 5 (AS), Ceramide 6 (NH), Ceramide 7 (AP),Ceramide 8 (AH), or Ceramide 9 (EOP).

In further embodiments, the invention includes a long-term use ofambroxol for improving skin barrier function, the method comprisingtopically administering a therapeutically effective amount of ambroxol,wherein the administration of the compound is at least for 1, 2, 3, 4,5, 6, 7, 8, 9, or 10 years. Administration of ambroxol could betopically applied several times over a given period of time, e.g., oncedaily, twice daily dose, three times daily.

In further preferred embodiments, the appearance of the skin (due to,for example, dryness, scarring, hyperpigmentation, etc) is improved, orpromoted, by up to 10%, 20%, 30%, 40%, 50%, 60%, or 70% as compared tountreated control subjects.

In preferred embodiments, the subject is a mammal, and even in furtherpreferred embodiments, the mammal is a human.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 shows the results of the time course of the return of TEWLtowards the baseline value with ambroxol in 100% dimethyl sulfoxide.

FIG. 2 shows the results of the time course of the return of TEWLtowards the baseline value with ambroxol in an alternative formulationof 60% dimethyl sulfoxide: 40% water [v/v].

FIG. 3 shows the effect of various ambroxol concentrations on TEWL,where the ambroxol concentration ranges from 20 uM to 2.0 mM.

FIG. 4 shows the 12 ceramide classes based on known sphingoid bases andfatty adds.

FIG. 5 shows the chemical structures and nomenclature for someceramides. Chemical structure and nomenclature of CER of the SC;A=α-Hydroxy fatty acid, EO=Ester-linked ω-Hydroxy acid, N=Nonhydroxyfatty acid, P=Phytosphingosine, S=Sphingosine andH=6-Hydroxysphingosine; Ceramide X=Nacyl dihydrosphingosine co-migrateswith ceramide 2 and is not included in this general analysis.

DETAILED DESCRIPTION

I. Definitions

The following definitions are provided for specific terms which are usedin the following written description.

As used in the specification and claims, the singular form “a”, “an” and“the” include plural references unless the context clearly dictatesotherwise. For example, the term “a cell” includes a plurality of cells,including mixtures thereof. The term “a nucleic acid molecule” includesa plurality of nucleic acid molecules.

The present invention can “comprise” (open ended) or “consistessentially of” the components of the present invention as well as otheringredients or elements described herein. As used herein, “comprising”means the elements recited, or their equivalent in structure orfunction, plus any other element or elements which are not recited. Theterms “having” and “including” are also to be construed as open endedunless the context suggests otherwise. As used herein, “consistingessentially of” means that the invention may include ingredients inaddition to those recited in the claim, but only if the additionalingredients do not materially alter the basic and novel characteristicsof the claimed invention.

As used herein, a “subject” is a vertebrate, preferably a mammal, morepreferably a human. Mammals include, but are not limited to, murines,simians, humans, farm animals, sport animals, and pets. In otherpreferred embodiments, the “subject” is a rodent (e.g., a guinea pig, ahamster, a rat, a mouse), murine (e.g., a mouse), canine (e.g., a dog),feline (e.g., a cat), equine (e.g., a horse), a primate, simian (e.g., amonkey or ape), a monkey (e.g., marmoset, baboon), or an ape (e.g.,gorilla, chimpanzee, orangutan, gibbon). In other embodiments, non-humanmammals, especially mammals that are conventionally used as models fordemonstrating therapeutic efficacy in humans (e.g., murine, primate,porcine, canine, or rabbit animals) may be employed. In preferredembodiments, an “individual” or “patient” (as in the subject of thetreatment) means mammals, particularly non-human primates, e.g. apes andmonkeys, and most particularly humans.

As understood herein, an “effective amount” of a topical composition ofthe instant invention refers to an amount of the composition suitable toelicit a beneficial response in the subject, e.g., for improving skinbarrier function, such as for example, increasing skin hydration, CERproduction, and/or healthy aging of the skin and/or to decrease TEWL ina subject.

The term “dose” or “dosage” as used herein refers to physically discreteunits suitable for administration to a subject, each dosage containing apredetermined quantity of the active pharmaceutical ingredientcalculated to produce a desired response.

The term “about” or “approximately” means within an acceptable range forthe particular value as determined by one of ordinary skill in the art,which will depend in part on how the value is measured or determined,e.g., the limitations of the measurement system. For example, “about”can mean a range of up to 20%, preferably up to 10%, more preferably upto 5%, and more preferably still up to 1% of a given value.Alternatively, particularly with respect to biological systems orprocesses, the term can mean within an order of magnitude, preferablywithin 5-fold, and more preferably within 2 fold, of a value. Unlessotherwise stated, the term ‘about’ means within an acceptable errorrange for the particular value, such as ±1-20%, preferably ±1-10% andmore preferably ±1-5%. In even further embodiments, “about” should beunderstood to mean +/−5%.

Where a range of values is provided, it is understood that eachintervening value, between the upper and lower limit of that range andany other stated or intervening value in that stated range isencompassed within the invention. The upper and lower limits of thesesmaller ranges may independently be included in the smaller ranges, andare also encompassed within the invention, subject to any specificallyexcluded limit in the stated range. Where the stated range includes oneor both of the limits, ranges excluding either or both of those includedlimits are also included in the invention.

All percentages and ratios used herein are by weight of the totalcomposition unless otherwise indicated herein. All temperatures are indegrees Celsius unless specified otherwise. All measurements made are atan ambient air temperature of 18-25° C., at normal atmospheric pressure,and at a relative humidity of 30-50%, unless otherwise designated.

All ranges recited herein include the endpoints, including those thatrecite a range “between” two values. Terms such as “about,” “generally,”“substantially,” “approximately” and the like are to be construed asmodifying a term or value such that it is not an absolute, but does notread on the prior art. Such terms will be defined by the circumstancesand the terms that they modify as those terms are understood by those ofskill in the art. This includes, at very least, the degree of expectedexperimental error, technique error and instrument error for a giventechnique used to measure a value.

Where used herein, the term “and/or” when used in a list of two or moreitems means that any one of the listed characteristics can be present,or any combination of two or more of the listed characteristics can bepresent. For example, if a composition of the instant invention isdescribed as containing characteristics A, B, and/or C, the compositioncan contain A feature alone; B alone; C alone; A and B in combination; Aand C in combination; B and C in combination; or A, B, and C incombination.

The term “isolated compound” means a compound substantially free ofcontaminants or cell components with which the compound naturallyoccurs, or the reagents used in synthesis or the byproducts ofsynthesis. “Isolated” and “substantially free of contaminants” does notmean that the preparation is technically pure (homogeneous), but it issufficiently pure to provide the compound in a form in which it can beused therapeutically.

As used herein, “skin barrier function” or a “skin's barrier function”is used consistently with its usage in the art—mainly to describe thefunction of the skin (which includes hair) to protect a subject fromexternal hazards, including microbial, chemical and/or mechanicalinsults. As used herein, the skin barrier function can be measured bynumerous ways, including: (a) the amount of time needed to recover theskin barrier function after an insult; (b) the amount of transepidermalwater loss (TEWL); (c) the amount of electrolyte loss; (d) the amountand/or healing time of dry skin; (e) the amount and/or healing time ofpruritus; (f) the amount and/or healing time of skin lesions; (g) theamount of skin hydration; (h) the ability to protect a subject fromforeign insults (e.g., antimicrobial barrier); (i) the amount and/orefficiency of ceramide (CER) and/or glucosyl-CER production, preferablyin the stratum basale, stratum pinosum, stratum granulosum, and/orstratum corneum layers of the skin; (j) the amount and/or efficiency inconverting glucosyl-CERs into CER, preferably in the stratum granulosumand stratum corneum layers of the skin; (k) the amount and/orconcentration of CER in the skin, preferably in the stratum corneumlayer of the skin; and/or (l) glucocerebrosidase (GCE) activity in thestratum basale, stratum spinosum, stratum granulosum, and/or stratumcorneum. Methods for measuring skin barrier function are well known andwell described in the literature. See, for example, Antonov et al.,“Methods for the Assessment of Barrier Function,” Curr Probl Dermatol.Basel, Karger, 2016, vol 49, pp 61-70; Feingold and Elias “Role ofLipids in the Formation and Maintenance of the Cutaneous PermeabilityBarrier” Biochimica et Biophysica Acta 1841:280-294 (2014); Breiden andSandhoff, “The Role of Sphingolipid Metabolism in Cutaneous PermeabilityBarrier Formation,” Biochimica et Biophysica Acta 1841:441-452 (2014) aswell as the references cited therein, all of which are incorporated byreference in their entirety.

Besides the biochemical measures used to evaluate “improved skin barrierfunction”, topical administration of ambroxol as described herein can bemeasured visually, such as, for example, by improved cosmetic signs ofaging or the means of reducing the risk of occurrence, delaying theonset, slowing the progression, and/or reducing the severity and/ormanifestation, of a sign of aging and/or degeneration of the skin, andincluding, but not limited to, preventing the occurrence, development orprogression of a sign of aging and/or degeneration of the skin. Otherpreferred embodiments include, decreasing the aging process of the skin,reducing the appearance of lines (e.g., fine lines) and/or wrinkles,decreasing skin sagging and/or imperfections, and/or improving and/orpromoting skin elasticity or suppleness, and/or increasing and/orpromoting retention of fluid in the skin. In other embodiments, animproved skin barrier function can be demonstrated by reducing sundamage (particularly UV radiation-induced oxidative stress/photodamage),stretch marks, and/or scars, reducing the appearance of burns, reducingthe appearance of blemishes (e.g. acne), reducing the appearance of darkcircles, age spots, and other hyperpigmentation, reducing skinthickness, loss of elasticity and/or collagen content, reducing dry skinand/or puffiness, preventing water loss of the skin, and/or increasingmoisturization, and/or reducing lentigines, freckles, and/or melasmas.

As used herein, “autophagy” describes the fundamental process fordegrading and recycling cellular components. During autophagy, damaged,unnecessary, dysfunctional macromolecules and organelles are broken downand are recycled for building new cellular components. In particular,autophagy makes it possible to regulate, repair and eliminate proteinswith a long service life in the cells, thus promoting cellularhomeostasis during differentiation and aging of human skin. Autophagy isthought to play a role in skin care. Moreover, ambroxol has been shownto modulate autophagy. See, for example PCT/US2018/042193 (hereinincorporated by reference in its entirety.)

As used herein, “ceramide” refers to a molecule composed of a sphingoidbase conjugated to a fatty acid via an amide bond and includes the 12different subclasses (shown in FIG. 4 ); at least 11 of these 12subclasses are known to be associated with human skin. Ceramides areprominent lipids found in stratum corneum, the outermost layer ofepidermis, and have an important function in the formation and retentionof the skin barrier in the stratum corneum. Pseudoceramides have similarstructures to natural ceramides. Skin damage caused by detergents whichremove the lipids essential for the barrier function will result in anincreased transdermal water loss (TEWL), and deteriorated barrierfunction has negative consequences for the total condition of the skin.A damaged skin barrier leads to increased skin sensitivity and potentialirritation such as atopic dermatitis or psoriasis. Topical applicationsof ceramide or pseudoceramide containing compositions are effective inrelieving atopic eczema. Ceramide or pseudo-ceramide also exhibittherapeutic properties such as wound and ulcer healing through thepromotion of cell restoration and growth.

Over 340 different species of ceramides have been classified based onthe composition of the head group or esterification of the fatty acid.Currently there are four different sphingoid bases and 3 different typesof fatty acids. FIG. 5 outlines the chemical structures and nomenclatureof the different classes of ceramides. See, also, Farwick et al.“Developments in Ceramide Identification, Synthesis, Function andNomenclature,” Cosmetics & Toiletries, Vol 124, No. 2 (2009). As usedherein, preferred ceramides used in combination with ambroxol, include,but are not limited to ceramides selected from: Ceramide EOS, CeramideNS, Ceramide NP, Ceramide EOH, Ceramide AS, Ceramide NH, Ceramide AP,Ceramide AH, Ceramide EOP, Ceramide NdS, Ceramide AdS, and/or CeramideEOdS.

As used herein, the phrase, “cosmetically acceptable delivery vehicle”is used to describe the components of the formulation other than theactive ingredients of the ambroxol formulation that are generallyrecognized as safe and nontoxic at the levels employed to achieve 100%weight of the formulation applied or delivered to the skin surface. Acosmetically acceptable vehicle may take the form of any known in theart that is suitable for application to skin. The vehicle may comprisean aqueous phase, an oil phase, an alcohol, a silicone phase or mixturesthereof. The cosmetically acceptable vehicle may also comprise anemulsion. Non-limiting examples of suitable cosmetically acceptablevehicles are disclosed in U.S. Pat. No. 9,238,000, entitled, “Method ofImproving Aging Appearance of Skin by Modulation of WIPI-1”; theteachings are incorporated herein by reference.

As used herein, a “cleanser” is formulated with ambroxol as describedherein to gently exfoliate dead skin cells, dissolve pore-cloggingimpurities, soothe and condition and prepare skin for the rest ofregimen. Examples of additional components that can be included in acleanser formulation include but are not limited to pomegranate extractto gently remove excess oil and dulling impurities to re-texturize skinand tighten appearance of pores and Tsubaki oil and rosehip oil toreduce inflammation and replenish essential lipids for improved textureand a comfortable after feel.

As used herein, an “essence” product is formulated with ambroxol asdescribed herein to protect the skin from harmful environmentalstressors, rehydrate, soothe and condition the skin. Additionalcomponents that can be added to an essence include but are not limitedto green tea extract to achieve a pH-balancing mist that shields theskin from environmental pollutants and aggressors while sealing inhydration and soothing skin and/or orange peel oil to help detoxifyskin, while high potency antioxidants including polyphenols, white tea,vitamin B and vitamin C offer all day protection from free radicals. Inaddition, nourishing fruit extracts and hyaluronic acid can be added tothe delivery vehicle to revive dehydrated and dull skin, reducepuffiness and discourage the formation of wrinkles.

As used herein, a “serum product” formulated with ambroxol as describedherein promotes, inter alia, cellular regeneration, improves firmnessand elasticity, reduces discolorations, brightens complexion, enhancesskin tone and texture. Additional components of a serum formulation caninclude an oil/water emulsion to achieve a serum that gives back to theskin what it naturally loses over time and the ability to self-repair;probiotics and Vitamin D3 to encourage skin renewal and support collagenand elastin production, enhancing firmness and elasticity: licoriceroot, niacinamide and Vitamin C can be included in the delivery vehicleto reduce dark spots and brighten skin tone for a noticeably glowingcomplexion. The lightweight formula and ingredients adapt to the skin'sunique needs and reignite luminosity, elasticity and suppleness.

As used herein a “day cream” can be formulated with ambroxol to alsoinclude a UVA and UVB broad spectrum SPF 30 ingredient, reduce rednessand inflammation, prevent collagen breakdown from damaging freeradicals, provide all-day hydration and plump the skin. Other componentsthat can be added, include but are not limited to an oil/water emulsionto achieve a day cream that delivers a precise balance of hydration andprotection to actively combat the visible signs of aging. SPF 30 defendsagainst harmful ultraviolet (UV) rays and free radicals while fightingwrinkles and visibly plumping skin, such as for example, UVA and UVBbroad spectrum SPF 30 ingredients, and/or antioxidants to protect fromthe sun's damaging rays and neutralize free radicals, peptides and plantstem cells to improve collagen production to visibly lift sagging skinand prevent the onset of wrinkles. Hyaluronic acid and tremella mushroomcan also be added to the delivery vehicle to deeply hydrate for smooth,plump skin.

As used herein, a “night cream” is formulated with ambroxol to deeplymoisturize and balance natural oils, lift, firm and sculpt facialcontours, reduce depth of fine lines and wrinkles and revitalize dull,tired skin. Other components that can be added to a night creamformulation include but are not limited to a light oil/water emulsion toachieve a night cream that provides the skin with unique support neededto maximize repair and renewal during sleep. Such components include butare not limited to argan oil, Vitamin B5 and hyaluronic acid to hydrate,nourish and combat dry, dull skin and improve barrier function. Peptidesalong with Vitamins A and C can be included in the delivery vehicle toencourage collagen and elastin production improving firnmess anddiminishing the appearance of fine lines and wrinkles for smooth, plumpskin. The rich, whipped texture deeply nourishes skin and redefinesfacial contours overnight to produce a firm and glowing complexion whenwaking.

As used herein, a “booster” product is formulated with ambroxol to helpeffectively up-regulate the skin autophagy and/or skin barrier functionfor dramatically younger-acting skin, which can reduce, for example,visible lines while increasing skin elasticity. The delivery vehicle inbooster formulations can include Vitamin C, Vitamin D, Vitamin B3 andomega-6 oil to target and treat visible lines while increasing skinelasticity.

Compounds of the Invention

Ambroxol, also known by its chemical nametrans-4-(2-amino-3,5-dibromobenzylamino) cyclohexanol, has the structureas shown in Table 1. As used herein, reference to “ambroxol” throughoutthe specification includes all salts of ambroxol. One example of such asalt is the a hydrochloride salt also shown in Table 1.

TABLE 1 Ambroxol

Ambroxol Hydrochloride

The term “salts” embraces addition salts of free acids or free bases.The term “pharmaceutically-acceptable salt” refers to salts whichpossess toxicity profiles within a range that affords utility inpharmaceutical applications. Pharmaceutically unacceptable salts maynonetheless possess properties such as high crystallinity, which haveutility in the practice of the present invention, such as for exampleutility in process of synthesis, purification or formulation oftherapeutic compounds.

Suitable pharmaceutically-acceptable acid addition salts may be preparedfrom an inorganic acid or from an organic acid. Examples of inorganicacids include, but are not limited to, hydrochloric, hydrobromic,hydriodic, nitric, carbonic, sulfuric, and phosphoric acids. Appropriateorganic acids may be selected from aliphatic, cycloaliphatic, aromatic,araliphatic, heterocyclic, carboxylic and salfonic classes of organicacids, examples of which include formic, acetic, propionic, succinic,glycolic, gluconic, lactic, malic, tartaric, citric, ascorbic,glucuronic, maleic, fumaric, pyruvic, aspartic, glutamic, benzoic,anthranilic, 4-hydroxybenzoic, phenylacetic, mandelic, embonic (pamoic),methanesulfonic, ethanesulfonic, benzenesulfonic, pantothenic,trifluoromethanesulfonic, 2-hydroxyethanesulfonic, p-toluenesulfonic,sulfanilic, cyclohexylaminosulfonic, stearic, alginic, s-hydroxybutyric,salicylic, galactaric, oxalic, malonic and galacturonic acid. Examplesof pharmaceutically unacceptable acid addition salts include, forexample, perchlorates and tetrafluoroborates. All of these acid additionsalts maybe prepared from ambroxol by reacting, for example, theappropriate acid with the compound.

Suitable pharmaceutically acceptable base addition salts of ambroxolinclude, for example, metallic salts including alkali metal, alkalineearth metal and transition metal salts such as, for example, calcium,magnesium, potassium, sodium and zinc salts. Pharmaceutically acceptablebase addition salts also include organic salts made from basic aminessuch as, for example, N,N′-dibenzylethylenediamine, chloroprocaine,choline, diethanolamine, ethylenediamine, m eglumine (N-methylglucamine) and procaine. Examples of pharmaceutically unacceptable baseaddition salts include lithium salts and cyanate salts. All of thesebase addition salts may be prepared from ambroxol by reacting, forexample, the appropriate base with the compound.

Ambroxol, including its salts, are known and may be prepared by methodsknown to the person skilled in the art of organic synthesis. Forexample, U.S. Patent Application publication number US2004/0242700,incorporated herein by reference in its entirety, provides a syntheticprotocol for the preparation of ambroxol.

Topical Compositions

In an aspect, the invention includes a composition comprising atherapeutically effective amount of ambroxol, in conjunction with apharmaceutically acceptable excipient for topical administration toimprove skin barrier function. Ambroxol may be administered in the formof a topical composition, in combination with a pharmaceuticallyacceptable carrier.

Ambroxol has been used topically in patients for the treatment of pain.See, Kern and Weiser, “Topical Ambroxol for the Treatment of NeuropathicPain,” Der Schmerz, Suppl 3, 29: S89-96 (2015). Here, a topicalformulation comprising 20% ambroxol was tested on patients to determinewhether ambroxol would alleviate and/or eliminate pain. The authorsreported that the pain reduction at this concentration was between 33and 100% which they deemed clinically relevant. Importantly, the authorsreported that no skin changes were observed even though other casereports suggested ambroxol caused systemic contact dermatitis. See,Monzon et al., “Ambroxol-Induced Systemic Contact Dermatitis Confirmedby Positive Patch Test,” Allergol Immunolpathol 37:167-169. Thus, it wasknown in the literature before the filing of this invention thatambroxol either caused adverse skin changes or had no effect on theskin's appearance.

However, it is surprisingly unexpected that doses substantially lessthan 20% would in fact improve skin conditions. It is believed thattopical administration of ambroxol to improve skin barrier function canoccur at formulation concentrations of less than 0.1% (w/w), and morepreferably, less than 0.01% (w/w), and even more preferably between0.001%-0.01% (w/w), and even more preferably between 0.0001-0.001%(w/w), and even more preferably between 0.00001-0.0001% (w/w). Ranges offormulation concentrations, all less than 0.1% (w/w) as described hereinare also specifically contemplated. It is unexpected and not suggestedin the literature that ambroxol could improve skin barrier function atsuch low formulation concentrations.

“Pharmaceutically acceptable carrier” means any carrier, diluent orexcipient which is compatible with the other ingredients of theformulation and not deleterious to the recipient. The active agent maybe formulated into dosage forms according to standard practices in thefield of pharmaceutical preparations. See Alphonso Gennaro, ed.,Remington's Pharmaceutical Sciences, 18th Edition (1990), MackPublishing Co., Easton, Pa. For examples of the preparation of ambroxolis disclosed in, for example, Examples 1-8 of WO2005/007146, or itsequivalent US2005/00148747, incorporated herein by reference.

For topical administration, ambroxol may be mixed with a suitablecarrier or diluent such as water, an aqueous buffer, an oil(particularly a vegetable oil), ethanol, saline solution, aqueousdextrose (glucose) and related sugar solutions, glycerol, or a glycolsuch as propylene glycol or polyethylene glycol. Solutions for topicaladministration preferably contain a water-soluble salt of the activeagent. Stabilizing agents, antioxidant agents and preservatives may alsobe added. Suitable antioxidant agents include sulfite, ascorbic acid,citric acid and its salts, and sodium EDTA. Suitable preservativesinclude benzalkonium chloride, methyl- or propyl-paraben, andchlorbutanol. The composition for topical administration may take theform of an aqueous or non-aqueous solution, dispersion, suspension oremulsion.

In preferred embodiments, ambroxol is formulated with a solvent thatenables ambroxol to permeate the upper layers of the skin. In preferredembodiments, the solvent is an organic solvent, such as for example,dimethyl sulfoxide.

Suitable permeation promoters according to the invention are urea,dimethyl sulfoxide (DMSO), hyaluronic acid sodium salt, alkanols such aslaurylalcohol or oleylalcohol, alkanoic acids such as oleic acid,1-dodecylazacycloheptan-2-one, ethyleneglycol, propyleneglycol ormenthol, as well as other permeation promoters selected from among the1-acylglycosides, 1-acyl-polyoxyethylenes, 1-acyl-saccharides,2-n-acyl-cyclohexanones, 2-n-acyl-1,3-dioxolanes (SEPA),1,2,3-triacyl-glycerols, 1-alkanols, 1-alkanoic acids, 1-alkyl-acetates,1-alkyl-amines, 1-alkyl-n-alkyl-polyoxyethylenes, 1-alkyl-alkylates,n-alkyl-beta-D-thioglycosides, 1-alkyl-glycerides,1-alkyl-propyleneglycols, 1-alkyl-polyoxyethylenes,1-alkyl-2-pyrrolidones, alkyl-acetoacetates, alkyleneglycols,alkylmethylsulphoxides, alkyl-propionates, alkylsulphates,diacylsuccinates, diacyl-N,N-dimethylaminoacetates (DDAA),diacyl-N,N-dimethylaminoisopropionates (DDAIP) and phenylalkylamines.

Topical formulations may also be combined with at least one excipientsuch as fillers, binders, humectants, disintegrating agents, solutionretarders, absorption accelerators, wetting agents, absorbents orlubricating agents.

Topical compositions comprising low dose formulations of ambroxol mayfurther comprise ceramides, which are well known in the art. See, forexample, Meckfessel and Brandt, “The Structure Function and Importanceof Ceramides in Skin and Their Use as Therapeutic Agents in Skin-CareProducts,” J Am Acad Dermatol 2014; 71:177-84 (herein incorporated byreference in its entirety.) In further preferred embodiments, thetopical formulation is used several times, giving doses over a period oftime, e.g., a daily dose or twice daily treatment for a week or more.

Examples of ceramides that can be formulated with ambroxol as describedherein include, but are not limited to: Ceramide EOS, Ceramide NS,Ceramide NP, Ceramide EOH, Ceramide AS, Ceramide NH, Ceramide AP,Ceramide AH, Ceramide EOP, Ceramide NdS, Ceramide AdS, and/or CeramideEOdS.

In further embodiments, the invention includes a long-term method of usein a subject, and/or to treat, inhibit, and/or reduce the skin agingprocess, skin age-related symptoms, and/or the skin age-relateddiseases, and/or to improve skin barrier function, preferably in ahuman, the method comprising administering a therapeutically effectiveamount of ambroxol, wherein the administration of the compound is atleast for 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 years.

In further preferred embodiments, skin barrier function is improved byup to about 10%, 20%, 30%, 40%, 50%, 60%, or 70% as compared tountreated control subjects. In further preferred embodiments, the skinbarrier function is improved by at least 10%, 20%, 30%, 40%, 50%, 60%,or 70% as compared to untreated control subjects. Ways to measure skinbarrier function are well known in the art and described herein.

The topical compositions of the present invention may also be formulatedso as to provide slow or controlled release of ambroxol therein using,for example, hydropropylmethylcellulose in varying proportions toprovide the desired release profile, other polymer matrices, gels,permeable membranes, osmotic systems, multilayer coatings,microparticles, liposomes and/or microspheres. In these embodiments,these formulations are ideally suited for improving skin barrierfunction.

In general, a controlled-release preparation is a topical compositioncapable of releasing ambroxol at the required rate to maintain constantpharmacological activity for a desirable period of time. Such dosageforms provide a supply of a drug to the body during a predeterminedperiod of time and thus maintain ambroxol levels in the therapeuticrange for longer periods of time than conventional non-controlledformulations.

The controlled-release of the active ingredient may be stimulated byvarious inducers, for example pH, temperature, enzymes, water, or otherphysiological conditions or compounds. Various mechanisms of drugrelease exist. For example, in one embodiment, the controlled-releasecomponent may swell and form porous openings large enough to release theactive ingredient after administration to a patient. The term“controlled-release component” in the context of the present inventionis defined herein as a compound or compounds, such as polymers, polymermatrices, gels, permeable membranes, liposomes and/or microspheres, thatfacilitate the controlled-release of the ambroxol in the topicalcomposition. In another embodiment, the controlled-release component isbiodegradable, induced by exposure to the aqueous environment, pH,temperature, or enzymes in the body. In another embodiment, sol-gels maybe used, wherein the active ingredient is incorporated into a sol-gelmatrix that is a solid at room temperature. This matrix is placed on theskin having a body temperature high enough to induce gel formation ofthe sol-gel matrix, thereby releasing ambroxol.

Without further description, it is believed that one of ordinary skillin the art can, using the preceding description and the followingillustrative examples, make and utilize the compounds of the presentinvention and practice the claimed methods. The following examples,therefore, specifically point out the preferred embodiments of thepresent invention, and are not to be construed as limiting in any waythe remainder of the disclosure. Although the invention herein has beendescribed with reference to embodiments, it is to be understood thatthese embodiments, and examples provided herein, are merely illustrativeof the principles and applications of the present invention. It istherefore to be understood that numerous modifications can be made tothe illustrative embodiments and examples, and that other arrangementscan be devised without departing from the spirit and scope of thepresent invention as defined by the appended claims. All patentapplications, patents, literature and references cited herein are herebyincorporated by reference in their entirety.

EXAMPLES Example 1. Restoration of Skin Barrier Function afterDisruption by Dimethyl Sulfoxide

Dimethyl sulfoxide (DMSO) is a well-known skin permeability enhancerthat is thought to act by disrupting ceramide-containing lipid bilayersof the stratum corneum and thereby compromising the permeability barrierof the skin (Agner and Serup, Clin. Expt. Dermatology 14:214 [1989];Notman et al Biophysical Journal 93:2056 [2007]). Among its manyprotective functions, this lipid barrier regulates transepidermal waterloss (TEWL) and is vital for the prevention of skin dryness in mammals.Thus, it is accepted by researchers in the field, that measurements ofTEWL can be used to directly assay the integrity of the skinpermeability barrier (Fluhr et al Experimental Dermatology 15:483[2006]).

To test ambroxol's ability to improve TEWL (i.e., reduce the rate ofwater loss), Ambroxol-HCl (MW 414.6, EP grade, Shilpa Medicare Ltd.,Raichur, Karnataka—584 102, India) was dissolved to a concentration of2.0 mM (0.075% solution [w/w]) in 99.9% pure dimethyl sulfoxide (EPgrade, Heiltropfen, Germany). 20 microliter aliquots of the 2.0 mMambroxol solution in DMSO as well as the 99.9% pure DMSO were spreadonto patches of human (adult male) skin in the upper thigh area; thearea of skin covered by the solutions in each case was approximately 2cm². All experiments were conducted in a narrow range of relativehumidity (30-40%) and at stable room temperatures of 18-22° C.

Transepidermal water loss (TEWL) measurements, in units of g/m²-h, weretaken at various time points after applying the solutions (at t=0) tothe skin sites. TEWL values were assessed using a wirelessclosed-chamber system (VapoMeter, Delphin Technologies Ltd., Kuopio,Finland) that automatically calculates water evaporation rates over asmall area (˜1.3 cm²); each measurement took about 10 s (Fluhr et alExperimental Dermatology 15:483 [2006]). Prior to the beginning of theexperiment, and at the conclusion of the experiment, TEWL measurementsof adjacent untreated skin areas were recorded; the baseline TEWL ofuntreated skin was found to not change significantly during the courseof the experiment. All time points were measured in duplicate ortriplicate. FIG. 1 shows the results of this experiment. Both the“control” (DMSO only) curve and the “2 mM ambroxol” (DMSO containing 2.0mM ambroxol) curve initially show a slow trend back towards the baseline(pre-treatment) TEWL, reflecting the natural post-disruptionreestablishment of the skin barrier (Agner and Serup, Clin. Expt.Dermatology 14:214 [1989]). However, at about 60 minutespost-application, the ambroxol curve exhibited a sudden drop in TEWL,indicating a rapid burst of skin barrier healing over a period ofapproximately 30 minutes. The improvement in TEWL, a difference of 4-6g/m²-h between the two curves, then maintains itself over the remainingcourse of the experiment. This indicates that the burst of improvementin the transepidermal water loss between 60-90 minutes post-applicationled to a durable improvement in the skin barrier. [Symbols: Solid line,DMSO (control with no ambroxol); dot-dashed line, 2.0 mM ambroxol inDMSO; dotted line, untreated skin from areas adjacent to the treatmentsites. Shown for each point are the error bars (standard deviation ofduplicate or triplicate measurements); for some points the error barsare less visible because they were smaller than the diameter of thesymbol representing that point.]

Example 2. Restoration of Skin Barrier Function after Disruption by aDimethyl Sulfoxide-Containing Aqueous Formulation

An alternative skin disruption agent, involving an aqueous formulation,was tested to see whether this might affect the skin barrier restorationproperties of ambroxol. Accordingly, a 2.0 mM solution of ambroxol wasprepared using 60% DMSO (60:40 DMSO:water [v/v]) mixture as the solvent.60% DMSO has been reported to be the minimum concentration of DMSOneeded to disrupt ceramide-containing lipid bilayers in skin (Notman etal Biophysical Journal 93:2056 [2007]).

Experimental conditions were essentially identical to those used inExample 1, except 10 microliter aliquots of the 2.0 mM ambroxol solutionin DMSO:water (60:40 [v/v]), or the DMSO:water solution alone as acontrol, were spread onto patches of human skin in the upper thigh area.Results are shown in FIG. 2 . All time points were measured in duplicateor triplicate.

Like the experiment shown in FIG. 1 , both the control and theambroxol-treated skin exhibit a slow approach to the baseline TEWL levelover a period of hours. Also like in the experiment shown in FIG. 1 ,there is an enduring improvement of approximately 5 g/m²-h in the rateof water loss of the ambroxol-treated skin relative to skin treated withthe DMSO:water formulation alone. However, in this case the burst occursearlier in the time course, in the first hour post-application (forpractical reasons the earliest time point in this series was about 40minutes post-application, and the burst had already started at thatpoint). This indicates that by altering the ambroxol formulation thekinetics of skin barrier improvement can be modulated. [Symbols: Solidline, 60:40 (v/v) DMSO:water (control with no ambroxol); dot-dashedline, 2.0 mM ambroxol in 60:40 (v/v) DMSO:water; dotted line, untreatedskin from areas adjacent to the treatment sites. Shown for each pointare the error bars (standard deviation of duplicate or triplicatemeasurements); for some points the error bars are less visible becausethey were smaller than the diameter of the symbol representing thatpoint.]

Example 3. Effect of Ambroxol Dosage on TEWL

FIG. 3 shows a dose-ranging study testing ambroxol concentrations of 20micromolar, 60 micromolar, 0.2 mM, 0.6 mM, and 2.0 mM. Apart from theseadditional concentrations, conditions and the formulation (99.9% DMSO)were essentially the same as those described in Example 1, except inthis experiment 10 microliter aliquots were spread and the skin regiontested was on the volar forearm.

The data in FIG. 3 are presented as histograms, with only start points(at 1 hr post-application) and end-points (at 10 hr post application)shown. Two things are notable about this experiment. One, in contrast tothe other experiments, the control skin treated with DMSO alone did notregress to the baseline during the 10 hour time-window of datacollection, as was observed with the experiments in Examples 1 and 2; infact, the TEWL seemed to get slightly worse during that time period.This may reflect the relative delicacy of volar forearm skin compared toskin of the upper thigh, such that the same treatment may be more orless harsh depending on the skin area treated. The second notable resultfrom this experiment was that even concentrations of ambroxol much lowerthan 2.0 mM yielded positive results in terms of ameliorating theincrease in TEWL caused by the DMSO insult to the skin barrier. In fact,sub-millimolar concentrations of ambroxol may even be optimal for skincare in some contexts. [Symbols: Open rectangles, skin TEWL measured at1 hr post-application; hatched rectangles, skin TEWL measured at 10 hrpost-application; dashed line, average TEWL value (7.8±1.1 g/m²-hr) ofuntreated skin from areas adjacent to the treatment sites. Allmeasurements were taken in quadruplicate (n=4) or quintuplicate (n=5).Shown for each value are the error bars (standard deviation ofquadruplicate or quintuplicate measurements.]

The disclosures of each and every patent, patent application, andpublication cited herein are hereby incorporated herein by reference intheir entirety.

While the invention has been disclosed with reference to specificembodiments, it is apparent that other embodiments and variations ofthis invention may be devised by others skilled in the art withoutdeparting from the true spirit and scope of the invention. The appendedclaims are intended to be construed to include all such embodiments andequivalent variations.

What is claimed:
 1. A method of treating a subject suffering frompsoriasis or atopic dermatitis in a subject, wherein said methodcomprises topically administering a composition comprising a low doseformulation of ambroxol, wherein said low dose formulation comprisesless than 0.1% (w/w) of ambroxol.
 2. The method of claim 1, wherein themethod improves skin barrier function, wherein said skin barrierfunction is measured by: 1) a reduction in the signs of aging (orpremature aging), 2) a decrease in the aging process of the skin, 3) areduction in the appearance of lines and/or wrinkles, 4) a decrease inskin sagging, 5) an improvement and/or promotion in skin elasticity orsuppleness, 6) a reduction in sun damage, 7) a reduction in stretchmarks, 8) a reduction in the appearance of scars, 9) a reduction in theappearance of burns, 10) a reduction in the appearance of blemishes(e.g. acne), 11) a reduction in the appearance of dark circles, 12) areduction in the appearance of age spots, 13) a reduction in theappearance of puffiness, 14) a prevention in water loss of the skin,and/or 15) an increase in moisturization.
 3. The method according toclaim 2, wherein the skin barrier function is improved by up to about10%, 20%, 30%, 40%, 50%, 60%, or 70% as compared to untreated controlsubjects.
 4. The method of claim 2, wherein the skin barrier function isimproved by: a. a quickening of the recovery of the skin barrierfunction after an insult; b. a decrease in the delay in the skin barrierrecovery; c. a decrease or prevention in transepidermal water loss(TEWL); d. a decrease or prevention of electrolyte loss in the skin; e.a decrease or prevention of dry skin; f. a decrease or prevention ofpruritus; g. a decrease or prevention of skin lesions; h. an increase orimprovement in skin hydration; i. an improvement in protection fromforeign insults (e.g., antimicrobial barrier); j. an increase inceramide (CER) and/or glucosyl-ceramide production, preferably in thestratum basale, stratum spinosum, stratum granulosum, and/or stratumcorneum layers of the skin; k. an increase in conversion ofglucosyl-CERs into CER, preferably in the stratum granulosum and stratumcorneum layers of the skin; l. an increase in the amount and/orconcentration of CER in the skin, preferably in the stratum corneumlayer of the skin; m. an increase in glucocerebrosidase activity in thestratum basale, stratum spinosum, stratum granulosum, and/or stratumcorneum; and/or n. a decrease in the overall rate of loss of robustnessof the skin upon aging, according to generally accepted functionalmetrics that monitor such a decline; wherein an increase or decrease ofany one of (a)-(n) at least 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%,55%, 60%, or 65% as compared to the untreated state.
 5. The method ofclaim 4, wherein the concentration of CER that increased in the stratumcorneum is at least 10%, 20%, 30%, 40%, 50% or 60% higher as whencompared to the concentrations of cholesterol and/or free fatty acids orother suitable benchmarks.
 6. The method of claim 2, wherein theimproved skin barrier function is measured by increasing and/orimproving skin appearance, skin elasticity, healthy skin aging and/orskin moisture in a subject.
 7. The method of claim 2, wherein theimproved skin barrier function is measured by inhibiting, and/orreducing skin aging, a skin age-related symptom, and/or a skinage-related disease in the subject.
 8. The method of claim 1, whereinthe method decreases/prevents transepidermal water loss (TEWL).
 9. Themethod of claim 1, wherein composition comprises ambroxolhydrocholoride.
 10. The method of claim 9, wherein the compositionfurther comprises a ceramide.
 11. The method of claim 10, wherein theceramide is selected from: Ceramide EOS, Ceramide NS, Ceramide NP,Ceramide EOH, Ceramide AS, Ceramide NH, Ceramide AP, Ceramide AH,Ceramide EOP, Ceramide NdS, Ceramide AdS, and/or Ceramide EOdS.
 12. Themethod of claim 1, wherein autophagy within the skin cells is alsoinduced.
 13. The method of claim 1, wherein composition is topicallyapplied several times over a given period of time, e.g., once daily,twice daily dose, three times daily.
 14. The method of claim 1, whereinthe composition is topically applied for at least 1, 2, 3, 4, 5, 6, 7,8, 9, or 10 years.
 15. The method of claim 1, wherein the low doseformulation concentration of ambroxol is: (a) less than 0.01% (w/w); (b)between 0.01%-0.09% (w/w); (c) between 0.001%-0.01% (w/w); (d) between0.0001-0.001% (w/w); (e) between 0.00001-0.0001% (w/w); (f) at about0.00001% (w/w), 0.00002% (w/w), 0.00003% (w/w), 0.00004% (w/w), 0.00005%(w/w), 0.00006% (w/w), 0.00007% (w/w), 0.00008% (w/w), 0.00009% (w/w),0.0001% (w/w), 0.0002% (w/w), 0.0003% (w/w), 0.0004% (w/w), 0.0005%(w/w), 0.0006% (w/w), 0.0007% (w/w), 0.0008% (w/w), 0.0009% (w/w),0.001% (w/w), 0.002% (w/w), 0.003% (w/w), 0.004% (w/w), 0.005% (w/w),0.006% (w/w), 0.007% (w/w), 0.008% (w/w), 0.009% (w/w), 0.01% (w/w),0.02% (w/w), 0.03% (w/w), 0.04% (w/w), 0.05% (w/w), 0.06% (w/w), 0.07%(w/w), 0.08% (w/w), or 0.09% (w/w).
 16. The method of claim 15, whereinsaid low dose formulation comprises less than 0.01% (w/w) of ambroxol.17. The method of claim 16, wherein autophagy is induced in the skincell.
 18. The method of claim 17, wherein the concentration of CER isincreased in the stratum corneum by at least 10%, 20%, 30%, 40%, 50% or60% higher than it was prior to treatment, relative to theconcentrations of cholesterol and/or free fatty acids and/or othersuitable benchmarks, such as the concentrations of reference moleculesin the stratum corneum that are known to be unaffected by the treatment.19. The method of claim 18, wherein the composition comprises ambroxolhydrocholoride.
 20. The method of claim 19, wherein the compositionfurther comprises a ceramide.
 21. The method of claim 20, wherein theceramide is selected from: Ceramide EOS, Ceramide NS, Ceramide NP,Ceramide EOH, Ceramide AS, Ceramide NH, Ceramide AP, Ceramide AH,Ceramide 9 (EOP), Ceramide NdS, Ceramide AdS, and/or Ceramide EOdS. 22.The method of claim 1, wherein the composition is formulated as alotion, spray, cream, shampoo, serum, emulsion, gel, oil, mask, essence,toner or paste.
 23. The method of claim 1, wherein the compositionfurther comprises essential fatty acid, preferably omega-3 or omega-6oil.